Annals of Botany 64, 533-544, 1989 533

Long-term Partitioning, Storage and Remobilizationof 14C Assimilated by Trifolium repens (cv. Blanca)

J. E. DANCKWERTS* and A. J. GORDONfThe Animal and Grassland Research Institute, Hurley, Maidenhead, Berks SL6 4LR, UK

Accepted: 15 May 1989

A B S T R A C T

The fourth fully expanded leaf on the main stolon of white clover plants was exposed to 14CO2. Thereafter,quantitative and fractional analysis of the partitioning, storage and remobilization after defoliation of the14C labelled assimilate was sequentially conducted over a 2- to 3-week period.

In undefoliated plants, most 14C reached its final destination within 24 h of feeding. Forty percent ofassimilated 14C was lost through respiration, while the rest was exported, predominantly to meristems, butalso to roots, stolons and leaves. The 14C initially translocated to meristems was subsequently recoveredin stolon and leaf tissue as the plants matured.

Approximately 10% of assimilated 14C was invested into long-term storage in roots and stolons. Thesereserves were remobilized after both partial and total defoliation, and a portion of the remobilized 14C wasincorporated into new growth. Partly defoliated plants regrew more rapidly than totally defoliated plants,but more 14C reserve depletion took place in the totally defoliated treatment. Reserve depletion took placefrom both stolons and roots, but stolon reserves were preferentially utilized. Both high and low molecularweight storage compounds were involved.

Key words: Trifolium repens, white clover, assimilate partitioning, storage, remobilization, defoliation.

I N T R O D U C T I O N

High nitrogen fertilization costs associated withgraminaceous forage crops have resulted in in-creased attention being given to forage legumes,particularly with regard to their use in grass/legume mixtures. White clover is perhaps themost important forage legume in the UnitedKingdom, and aspects of its carbon and nitrogeneconomy have received increasing attention in theliterature. The partitioning of 14C during the 24 hfollowing assimilation by leaves of various ages isreported in detail by Ryle, Powell and Gordon(1981 a, b). This work did not, however, fractionatebetween structural and labile carbon, or considerlong-term storage and re-allocation of storedcarbon after defoliation.

In graminaceous plants, most assimilated carbonis either lost through respiration or metabolizedinto structural carbon or protein within a few days(Gordon, Ryle and Powell, 1977). A relatively

* Present addresses: Department of Agriculture(Eastern Cape Region), Private Bag XI5, Stutterheim,South Africa.

t Welsh Plant Breeding Station, Plas Gogerddan,Aberystwyth, Dyfed, SY23 3EB, UK.

small proportion of assimilate may be set aside forlong-term storage to be remobilized at a laterstage, should there be an inadequate supply ofphotosynthate. Numerous authors have stressedthe importance of reserve carbon in regrowth afterdefoliation (e.g. Hyder and Sneva, 1959; Wein-mann, 1961; Trlica, 1977; Daer and Willard,1981) but, in some instances, the traditional ideathat grasses deposit reserve carbon to be remobil-ized at a later date has been disputed (Ryle andPowell, 1974; Atkinson and Farrar, 1983;Richards and Caldwell, 1985). Danckwerts andGordon (1987) used 14C labelling techniques toshow that, although a relatively small fraction ofphotosynthate was invested in long-term labilereserves in Lolium perenne, this small fractioncould build up to form a large pool over anextended period. After defoliation, the reserveswere at least partly depleted and incorporated intonew growth.

Little published information exists regardinglong-term partitioning, storage and re-use ofassimilated carbon in forage legumes. This paper,therefore, reports on an investigation using acombination of l4C tracer techniques and chemicalanalyses (Gordon et al, 1977; Atkinson and

0305-7364/89/110533+ 12 $03.00/0 © 1989 Annals of Botany Company

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534 Danckwerts and Gordon—14C Assimilation in Trifolium repens

Farrar, 1983) to obtain a fractional and dynamicaccount of the fate of 14C incorporated throughsingle leaves of white clover plants {Trifoliumrepens L. cv. Blanca). The specific objectives of thestudy were: (i) to provide a long-term account ofthe partitioning of assimilated l4C in plant organs,(ii) to establish whether a fraction of assimilated14C was stored in a re-usable chemical form inroots and stolons, and, if so, (iii) to determinewhether storage 14C was remobilized and in-corporated into new growth after partial andcomplete defoliation.

MATERIALS AND METHODS

Plant material and growth conditions

White clover plants were cultured from clonalstolon tips and transferred singly into 100 mmdia. x 120 mm deep plots filled with Perlite. Theywere grown in a Saxcil growth cabinet providing a12 h photoperiod from a mixture of fluorescenttubes and incandescent lamps (630 /iEm'2 s"1 inthe spectral region 400-700 nm). The relativehumidity was maintained at about 60% and CO2

was injected to ensure the concentration did notfall below 300 /tl I"1.

After transfer of stolon tips, pots were inoculatedwith Rothamsted Rhizobium 221 and, thereafter,watered daily with no-nitrogen nutrient solution.This was supplemented with an additional ap-plication of demineralized water as the plants'water requirements increased with increasing ma-turity.

Experimental plants were selected for uniformityonce at least six fully expanded leaves and two tofour secondary stolons had been subtended by theprimary stolon.

I 4 C0 2 uptake

The fourth fully expanded leaf on the mainstolon of each experimental plant was fed with14CO2 for 30 min approx. midway throughthe photoperiod. This leaf was selected for feedingto ensure that cell division and expansion wascomplete and, therefore, that the fed leaf wouldnot be a major sink of photosynthate. Theapparatus and technique used to present l4CO2 tosingle leaves has been described elsewhere (Ryleand Powell, 1974, 1976; Ryle et al., 1981a).Petioles of fed leaves were marked with a spot ofdye to allow their identification at a later stage.

Treatments and sampling programme

Two separate trials were undertaken. The first(Expt 1) was aimed primarily at establishing long-

term partitioning of assimilated 14C and whether afraction of this carbon was set aside in storageorgans in a reusable form. The second trial (Expt2) aimed at testing whether stored labile carbonwas remobilized and incorporated into regrowthafter complete and partial defoliation.

Expt 1. Thirty plants were fed 14CO2 andsequentially harvested immediately after feedingand then 1, 4, 8, 11 and 18 d thereafter. Eachharvest consisted of five plants (replications). Plantswere harvested by washing Perlite away from rootswith minimum damage to plant material. Im-mediately after feeding, harvested plants wereseparated into two components, the fed leaf(including petiole) and the rest of the plant. At allother harvest dates plants were dissected into the

. fed leaf, roots stolons, meristems and all otherleaves (including petioles). In addition, on d 1 andd 4, the primary (fed) stolon and meristem wereseparated from other stolons and meristems.Meristems were defined as the expanding leaf andstolon material beyond the youngest full open leafon each stolon.

Expt 2. Thirty-two white clover plants weregrown and exposed to 14CO2 in an identical mannerto those in Expt 1. Four plants were harvestedimmediately after feeding and separated into thefed leaf and the rest of the plant. A further fourplants were harvested 4 d after feeding anddissected into roots, stolons, the fed leaf and allother material (leaves and meristems). On thesame day, the fed leaf was removed from allremaining plants, and the latter randomly allocatedinto three treatments; a control where plants werenot defoliated, a partial defoliation treatment anda total defoliation treatment. In the totallydefoliated treatment, all leaves and meristems wereremoved. The partly defoliated treatment involvedremoval of meristems and all but two leaves, thesebeing the leaf on the main stolon immediatelypreceding the fed leaf and the oldest fully expandedleaf on the second stolon.

Four plants from each treatment were harvested5 and 10 d after defoliation (9 and 14 d afterfeeding). At these harvests, control plants wereseparated into roots, stolons and the rest of theplant. Defoliated plants were separated into roots,stolons and new top-growth produced after de-foliation. Old leaves were separated from newgrowth in the partly defoliated treatment.

Extraction and analysis of plant samples

All plant parts harvested immediately afterfeeding were killed by immersion in boiling 80%(v/v) ethanol. Thereafter, only fed leaves andplant parts considered possible sites of long-term

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labile carbon storage that would still be availableafter defoliation (roots and stolons) were killed inethanol. An exception was in Expt 1 when primaryand secondary meristems were separated (1 and 4d after feeding). At these harvests, meristems, too,were killed in boiling ethanol. Otherwise, all plantsamples were dried to a constant mass at atemperature of 90 °C.

Complete ethanol extraction of relevant sampleswas achieved by repeated boiling and decantation.The ethanol insoluble material was dried, groundand the starch extracted as in MacRae (1971).Sub-samples of the ethanol and starch extractswere assayed for 14C content. A detailed de-scription of the chemical compounds contained inthese two extracts is provided by Gordon et al.(1977). In summary, however, these extracts wouldbe expected to contain virtually all the sources oflabile carbon (low and high molecular weightcompounds in ethanol and water extracts, re-spectively).

Plant residues remaining after extraction oflabile carbon as well as those plant parts notsubjected to extraction (or sub-samples of these)were burnt in a Packard Tri-carb oxidiser. Theabsolute radioactivity of these samples and that ofthe various extracts was determined using a Model460C Packard Tri-carb liquid scintillationcounter.

Analysis and presentation of data

Plant masses are presented as the means ofindividuals masses (whole plants) or as the meansof logarithms of individual values of plant parts.Transformation was undertaken to minimize vari-ance heterogeneity and to facilitate presentationof the masses of plant parts of high and low masson the same axes. In the case of whole plant mass,cubic regressions are fitted to the means. Thechoice of cubic functions is incidental, the purposeof curve fitting being merely to summarize the datain a convenient manner, and to facilitate derivationof continuous growth indices. The masses of thevarious plant parts are not regressed over timesince linking of means with straight lines ad-equately summarizes the results.

Radioactivity in the various plant fractions wasexpressed in kBq per plant part. Since the variationin the area of the exposed leaf was a major sourceof variation in total l4C per plant, radioactivitywas normalised to 1 m2 of leaf exposed to 14CO2.(The area of each fed leaf was recorded using alight interception planimeter.) The means of thesevalues are also not regressed over time and arelinked by straight lines or presented as histograms.Such presentation adequately summarized the

results, and in any event, the paucity of harvestdates precludes curve fitting in most instances.

Superimposed on all means presented in thefigures are twice the standard errors of the mean.These, therefore, represent an estimate of therange in which the true mean of the population ofplants is expected to have occurred.

RESULTS AND DISCUSSION

Plant mass

It is demonstrated in Fig. 1A that the pattern ofgrowth of plants was similar in both experiments,although the mass of the plants in Expt 1 was alittle larger than that of plants in Expt 2 at the timeof feeding with 14CO2. The absolute growth ratesof both populations of plants were increasing atthe time of feeding, and continued to do sothroughout the duration of both experiments.Relative growth rates (data not shown) increasedor remained constant during the first 8/9 d of eachexperiment. This implies that any storage of labile14C during this period would not have been theresult of a reduction in growth due to inadequatesupply of some other requirement for growth, suchas light or water.

The partitioned mass of undefoliated plants(Fig. 1 B) shows that by far the greatest portion ofthe plant consisted of leaves and not storageorgans such as roots or stolons.

It is clear also that defoliation resulted in adramatic drop in the production of roots andstolons. Total defoliation had a greater adverseeffect on root and stolon growth than partialdefoliation (Fig. 2 A). Furthermore, partly de-foliated plants produced more new growth afterdefoliation than totally defoliated plants (Fig. 2B).

Uptake, respiratory loss and partitioning of 14C inundefoliated plants

Total 14C declined by approximately 40 to 45 %during the first 24 h after feeding. This respiratoryloss was of a similar order to that reported byGordon et al. (1977) and Danckwerts and Gordon(1987) for uniculm barley and perennial ryegrass,respectively.

Radiocarbon assimilated by the fed leaf wasrapidly exported (Fig. 3B). Only 10% of total 14C.assimilated remained in the fed leaf after 24 h,declining to less than 5 % 3 d later. Other than thatlost through respiration, the largest portion of 14Cexported from the fed leaf within the first 24 h wastranslocated to aerial meristems. Somewhat lesswas exported to roots with relatively small amountsbeing found in stolons and leaves (excluding thefed leaf). This data is similar to that reported by

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536 Danckwerts and Gordon—liC Assimilation in Trifolium repens

18Days

FIG. 1. A, Total mass of plants in Expt 1 (Ml) and undefoliated plants in Expt 2 (M2) with derived absolute growthrates (AGR1 and AGR2, respectively); B, mass of plants in Expt 1 partitioned into leaves (L), meristems (M), stolons

(S) and roots (R). Bars represent twice standard errors of the means.

- I -

14

Days offer exposure H C 0 2

FIG. 2. A, Root (R) and stolon (S) mass of control (C), partly defoliated (Pd) and totally defoliated (Td) plants afterdefoliation in Expt 2; B, mass of new top growth after partial (Pd) and total (Td) defoliation. Bars represent twice

standard error of the means.

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10

X 8

8

I '

Perentage I4C depletion

Total radioactivity

50

40

30

S

20

10

0 I

3 -

18

Days after exposure to l4C02

18

FIG. 3. A, Total radioactivity found in whole plants and B, partitioned into the fed leaf (FL), other leaves (OL),meristems (M), stolons (S) and roots (R) following exposure of single leaves to I4CO2 for 30 min (Expt 1). Respiratoryloss of 14C, expressed as 'percentage 14C depletion' is also indicated in (A). Bars represent twice standard errors of

the means.

Ryle et al. (1981a and b) and merits no morediscussion here.

After d 1, the 14C in meristems itself declinedsharply and very little remained at the end of theexperimental period. The decline of 14C inmeristems was compensated for by an increase of14C in leaves and stolons (Fig. 3 B). This is probablydue to the fact that, as the plant grew, meristematictissue at early harvests had matured and becameeither leaf or stolon material at later harvests.

Total 14C in roots did not change significantlyafter d 1 (Fig. 3 B), supporting the contention thatthere was relatively little active translocation ofI4C between organs after d 1.

Partitioning of 14C between primary and sec-ondary stolons and meristems is presented in Fig.4. The primary meristem and stolon contained lessl4C than the secondary meristems and stolons,respectively, both 1 d and 4 d after feeding (Fig.4 A), indicating that stolons subtended by the sameplant do not act as independent units in whiteclover at this stage of growth.

The relatively high specific activity of meristems1 d after feeding (Fig. 4B) illustrates the highdemand made by meristems on the carbon

assimilated by mature leaves. The radioactivityand specific activity of meristems declined signifi-cantly 4 d after feeding (Fig. 4), a result of the l4Cin meristems being incorporated into leaves andstolons as the plant matured.

Distribution of 14C between labile and structuralfractions

Immediately after feeding, virtually all 14C in theplant was in a chemically labile form in the fedleaf. Distribution to other organs occurred rapidly(see above) and some of the l4C was convertedfrom labile to structural material in the process ofsynthesis of new tissue (Fig. 5). Between 1 d and 4d there was a decline in labile I4C in the fed leaf,meristems and roots. The decline in the fed leafcan be attributed to respiration and to trans-location of remaining labile 14C to other plantorgans. In the roots, there was a relatively smalldecline in labile 14C, and this can be accounted forby continued synthesis of structural material. Thedecline in labile 14C in the meristems is alsoattributed to synthesis of structural tissue, but by4 d after feeding much of this tissue was no longer

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S3t Danckwerts and Gordon—l4C Assimilation in Trifolium repens

12

S

10

PS

ss

SM

PSss

SM

I day after feeding 4 days after feeding I day after feeding 4 days after feeding

FIG. 4. A, Total radioactivity in the primary stolon (PS), primary meristem (PM), secondary stolons (SS) andsecondary meristems (SM) of undefoliated plants (Expt 1) 1 d and 4 d after feeding; B, specific activity of primaryand secondary stolons and meristems 1 d and 4 d after feeding. Bars represent twice standard errors of the means.

iFIG. 5. Radioactivity of structural (shaded) and labile (unshaded) 14C in the fed leaf (FL), meristems (M), stolons (S)and roots (R) of undefoliated plants (Expt 1) A, 1 d and B, 4 d after exposure to 14CO2. Bars represent twice standard

errors of the means.

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1-5

1-0

0-5

11 18 " I

Days after exposure l4C02

FIG. 6. A, Radioactivity of labile 14C in stolons, roots, and stolons and roots together of undefoliated plants (Exp1); B, radioactivity of combined labile 14C in stolons and roots expressed as a percentage of initial total 14C uptake

(1) and of current total 14C in the plant (2). Bars represent twice standard errors of the means.

in the meristem, and had developed into leaf orstolon material. Thus, the structural fraction of14C in stolons was able to increase between d 1 andd 4, without any decline in the stolon labile fraction(Fig. 5).

Long-term storage of labile carbon in undefoliatedplants

It is conceivable that labile carbon can be storedin virtually any plant organ. However, the objec-tives of the study were to consider those labilereserves that would be remobilizable after de-foliation. For this reason, only carbon stored inroots and stolons is considered. It is acknowledgedand emphasized that roots consist of both possiblestorage sites, and meristems. Root meristemswould not be expected to be sites of labile carbonstorage, but sinks for synthesis of new tissue.Nevertheless, a persistence of previously assimi-lated l4C in a labile form in roots and stolonswould indicate storage, even though it mayconstantly be drawn upon for synthesis of newtissue.

Labile 14C reserves in roots and stolons arepresented in Fig. 6. These (pooled) declined slightlybetween 1 d and 4 d after feeding, representing12-4 and 10-4 % of the original package ofassimilated 14C on these two respective days (Fig.6). After d 4, labile 14C reserves remained relativelyconstant (Fig. 6 A). Eighteen d after feeding,labile reserves in roots and stolons (pooled)represented approximately 19% of total 14Cremaining in the plant and about 10% of the totalinitially assimilated (Fig. 6B). This proportion isapprox. three times higher than the proportion oflabile 14C found in the roots and stem basesof perennial ryegrass 3 weeks after feeding(Danckwerts and Gordon, 1987).

The observed decline in labile 14C between d 1and d 4 (Fig. 6 A) was a result of a decline in theroots and not the stolons. The former includedroot meristems, and the drop was probably aresult of some of the 14C reserves being used to laydown new structural tissue. After d 4, the level ofroot reserves remained fairly constant, while stolonreserves, if anything, increased slightly. Theobserved increase in stolon reserves was perhaps a

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540 Danckwerts and Gordon—14C Assimilation in Trifolium repens

0-5

0-4

I 8

1 1 extract W

18I 18 IDays after exposure l4C02

FIG. 7. Radioactivity of labile 14C reserves in high molecular weight (water extract) and low molecular weight(ethanol extract) reserves in A, stolons and B, roots of undefoliated plants (Expt 1). Bars represent twice standard

errors of the means.

result of the fact that, as meristems developed intostoloniferous tissue, a portion of the labile l4Cinitially in the meristem that was not used tosynthesize structural tissue, remained in the newstolon in a labile form. Between 4 d and 18 d afterfeeding, labile 14C reserves in stolon exceededthose in roots (Fig. 6 A).

The labile 14C reserves in stolons and rootsconsisted of both high and low molecular weightcompounds (Fig. 7). However, the relative propor-tions of the two differed between the organs. Instolons, 14C in low molecular weight compoundsinitially exceeded those in high molecular weightcompounds, but the latter built up, and from 8 dafter feeding, exceeded 14C reserves in low mol-ecular weight compounds (Fig. 7 A). In roots,there was more l4C in low molecular weightreserves than in high molecular weight reserves,and the relative proportion between the two wasapproximately the same throughout the experi-mental period (Fig. 7B).

Rapid depletion of labile 14C assimilated byforage plants, and grasses in particular, has been

reported elsewhere (Gordon et al., 1977; Prosserand Farrar, 1981; Atkinson and Farrar, 1985).The rapid turnover rate of labile carbon has beenused to refute the idea that forage plants depositlong-term reserves to be remobilized at a later date(Atkinson and Farrar, 1983). In a study of long-term storage of labile 14C in perennial ryegrass,Danckwerts and Gordon (1987) also reportedrelatively rapid depletion of labile carbon. How-ever, they found that a relatively small proportionof assimilated carbon (4 %) was still present in alabile form in roots and stem bases 3 weeks later.They thus argued that this relatively small pro-portion of carbon invested in long-term reservescould build up to form a considerable pool over anextended period. In the current investigation, theproportion of assimilated carbon set aside forlong-term storage in roots and stolons in whiteclover (10% after nearly 3 weeks) was consider-ably higher than that reported by Danckwerts andGordon (1987) for perennial ryegrass.

If long-term carbon reserves are arbitrarilydefined as labile carbon that has been in roots and

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Danckwerts and Gordon—UC Assimilation in Trifolium repens 541

0-9

14 4 9

Days after exposure l4C

14

FIG. 8. Radioactivity of labile 14C reserves in partly defoliated (Pd), totally defoliated (Td) and control (C) plants inExpt 2. A, stolons and roots together; B, stolons; C, roots. Bars represent twice standard errors of means.

stolons for 4 d, and it is assumed, firstly, that thetrends in Fig. 6 are representative of carbon fromthe whole plant (not just from the fed leaf) and,secondly, that the plant is growing approximatelylinearly, then in an 18 d period the plant wouldaccumulate labile carbon reserves equal to nearlytwice the total carbon fixed by photosynthesisduring a single day at the mid-point of the 18 dperiod. It thus seems that white clover may havethe ability to build up a considerable pool ofstorage carbon available for remobilization fol-lowing an external perturbation such as defoli-ation.

Remobilization of storage carbon after defoliation

In plants clipped 4 d after exposure to 14CO2,labile 14C reserves were depleted far more rapidlythan in control plants (Fig. 8 A). The level of labilereserves in defoliated plants was significantly lowerthan that in control plants 5 d after clipping, andreserves were depleted further during the following5 d. There was no difference between the partly

and totally defoliated treatments in the amount oflabile 14C reserves depleted during the first 5 dafter clipping. However, during the second 5 dafter clipping, significantly more reserves weredepleted in the totally defoliated treatment than inthe partly defoliated treatment. Clearly, defoliationresulted in mobilization of a fraction of storagecarbon that did not take place in undefoliatedplants, and more reserves were mobilized aftertotal defoliation than after partial defoliation (Fig.8 A).

During the first 5 d after defoliation, mobiliz-ation of 14C reserves apparently took place fromstolons and not from roots (Fig. 8B, C). Duringthe second 5 d period after defoliation, stolonreserves continued to deplete slightly, but signifi-cant depletion of root reserves took place. Tendays after clipping, there was little difference in theamount of stolon reserve utilized between partialand total defoliation treatments. However, signifi-cantly more root reserves were used 10 d after totaldefoliation than after partial defoliation (Fig. 8B,C). It therefore seems that, after defoliation,

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542 Danckwerts and Gordon—14C Assimilation in Trifolium repens

0 - 2 5 -

>. 0 - 1 5 -

0-05 -

14 4

Days offer exposure to l4C02

14

FIG. 9. Radioactivity of A, low molecular weight reserves and B, high molecular weight reserves in stolons of partlydefoliated (Pd), totally defoliated (Td) and control (C) plants in Expt 2. Bars represent twice standard errors of means.

preferential use was made of stolon reserves and,after these had become depleted, the plant beganto use the root reserve pool. Greater demand wasmade on the reserve pool after total than afterpartial defoliation. Utilization of reserves tookplace for at least 10 d after clipping.

Both high and low molecular weight reserveswere remobilized (Fig. 9). In the stolons, there wasno significant difference in the level of ethanolextractable 14C utilized between the two utilizationintensities (Fig. 9 A). However, after partial de-foliation, somewhat less depletion of high mol-ecular weight reserves took place than after totaldefoliation (Fig. 9B). The implication is thatpreferential demand may be made for low mol-ecular weight carbon compounds after defoliation.

Utilization of remobilized carbon in new growthafter defoliation

It is shown in Fig. 10A that after defoliation, atleast some storage 14C was incorporated into newgrowth. This incorporation continued throughoutthe experimental period, although the plant had bythis stage produced new photosynthetically active

leaf material (Fig. 2). The amount of 14C reservesincorporated into new growth was significantlyhigher after total than after partial defoliation(Fig. 10A), despite the fact that the total amountof regrowth was greater after partial defoliation(Fig. 2).

The total depletion of labile l4C reserves fromstolons and roots (also shown in Fig. 10 A) wasconsiderably greater than the amount of 14Crecovered in new growth after clipping, showingthat some of the reserves depleted after clippingwere lost through respiration. The amount ofreserve carbon lost through respiration afterclipping was, in fact, greater than that incorporatedin new growth.

While 14C reserves may have been used to someextent for new growth and respiration in un-defoliated plants, because defoliation resulted invery much more rapid depletion of reserves thanno defoliation (Fig. 8), it is emphasized thatdefoliation does, indeed, result in remobilizationand use of reserve carbon that would not be usedin undisturbed white clover plants.

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9 14

Days after exposure to MCO2

FIG. 10. Accumulation of 14C in new growth (Ng) afterpartial (Pd) and total (Td) defoliation, and total depletion(depl.) of reserves from stolons and roots. Bars represent

twice standard errors of means.

CONCLUSION

Assimilate partitioning

The bulk of the 14CO2 assimilated by a mature, butviable leaf of an undisturbed white clover plantwas rapidly exported within 24 h. About 40 % waslost through respiration, while the remainder wasexported predominantly to meristems, with smallerproportions allocated to roots, stolons and leaves(other than the fed leaf)- After 24 h, 14C levels inroots remained relatively constant while a rapiddecline of 14C in meristems was accompanied by anincrease in leaves and stolons. This was probablynot a result of active translocation, but rather aresult of the fact that, as the plant grew, what hadpreviously been meristematic tissue became stolonand root material. It thus seems that most of theassimilated 14C had reached its final destinationwithin 24 h of feeding. About 4 % of totalassimilated l4C did not leave the fed leaf. A largeproportion of assimilated 14C was transportedfrom the fed to other stolons, indicating thabiologically-linked stolons do not act as inde-

pendent units. There was, however, a 14C con-centration gradient between the fed and otherstolons.

Although most 14C transport took place duringthe first 24 h after feeding, conversion of some ofthis 14C from labile compounds to structural tissuecontinued between 24 and 96 h after 14CO2 uptake.

Long-term carbon storage and remobilization

The level of total labile 14C reserves in stolonsand stem bases stabilized approx. 4 d after ex-posure to 14CO2. Root reserves declined between 1and 4 d after feeding, while stolon reservesstabilized (or increased slightly) 1 d after exposureto 14CO2. Eighteen d after feeding, 10% of the14C transported to the stolons and roots was stillpresent in these organs as labile reserves. If thisresult is representative of the whole plant, then aconsiderable pool of reserve carbon may be builtup over an extended period. Reserve 14C consistedof both low and high molecular weight compounds,but 18 d after feeding, stolons contained morehigh than low molecular weight reserves, while theopposite occurred in the roots.

Both partial and total defoliation resulted indepletion of labile reserves, but the depletion wasgreater after total than after partial defoliation.Reserves were remobilized from both stem basesand roots, but it seemed that stolon reserves werepreferentially utilized, and after these had beendepleted, root reserves were used. Both low andhigh molecular weight reserves were utilized,although there might have been some preferencefor low molecular weight compounds. At leastsome of the remobilized 14C was incorporated innew growth, although some reserve carbon wasused for respiration. More 14C was recovered inthe new growth of totally defoliated than in partlydefoliated plants, despite the fact that the mass ofthe new growth was greater after partial de-foliation.

Finally, the major conclusion drawn from theinvestigation is that white clover plants growing inundisturbed conditions do, indeed, invest a portionof current photosynthate in long-term labilereserves. After defoliation, the reserves are at leastpartly depleted, and some of the reserve carbon isincorporated in new growth. However, partialdefoliation results in more rapid regrowth and lessdepletion of reserves than total defoliation, imply-ing that concurrent photosynthesis also makes acontribution to new growth. The relative im-portance of reserve carbon versus concurrentphotosynthesis to regrowth after defoliation there-fore requires further investigation, as does theeffect of the size of the reserve pool.

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544 Danckwerts and Gordon—llC Assimilation in Trifolium repens

ACKNOWLEDGEMENTSWe acknowledge B.P. Southern Africa (Pty) Ltdwho funded the visit by the senior author to theAnimal and Grassland Research Institute, and thelatter Institute for providing facilities and fundingfor the research. We also thank Miss D. Mitchelland Mr C. E. Powell for technical assistance. TheAnimal and Grassland Research Institute isfinanced through the Agricultural and FoodResearch Council.

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